【Objective】To explore the inter and intraspecifically physiological and biochemical characteristics of three barrenwort species and to provide theoretical basis of standardized cultivation with high yield.【Method】 The morphology, the relative content of chlorophyll(SPAD value) and stomatal conductance of three transplanted barrenwort species were investigated. Non-parametric test and one-way ANOVA were used to analyze the data. 【Result and conclusion】 The order of plant height with young leaves from high to low was Epimedium wushanese Ying, Epimedium pubescen Maxim, Epimedium acuminatum Franch. And there were significant differences between E. acuminatum and the other two plants. The order of plant height with old leaves from high to low was E. wushanese, E. acuminatum, E. pubescen. And there were significant differences between E. pubescen and the other two plants. Significant differences of leaf morphology of plants with old leaves were observed among three species. But no significant differences was observed among plants with young leaves of three species. For three species, the chlorophyll contents of all plants with old leaves were higher than those of plants with young leaves. Significant differences of stomatal conductance were observed among three intraspecifically species with young leaves were slightly higher than those of old leaves. The different barrenwort species had different degress of adaptability to the growth environment.